Electrode arrangement for generating functional field barriers in microsystems

ABSTRACT

A microsystem adapted for dielectrophoretic manipulation of particles in a suspension liquid wherein the microsystem has a channel with channel walls and a longitudinal extension. An electrode arrangement is present which has at least one microelectrode on at least one of the channel walls. This acts to generate a field barrier which crosses the channel at least partly. The microelectrode has a band-shape or has a multitude of straight electrode sections connected to each other. The band-shape has a predetermined curvature or the straight electrode sections are arranged with predetermined different angles so that the field barrier has a predetermined curvature relative to the longitudinal extension of the channel.

This application is a 371 of PCT/EP99/04470, filed Jun. 28, 1999.

The invention relates to electrode arrangements for generatingfunctional field barriers in microsystems adapted for manipulation ofsuspended particles, in particular functional microelectrodes fordielectrophoretic deflection of microscopic particles, and microsystemsequipped with such electrode arrangements as well as their applications.

BACKGROUND OF THE INVENTION

Manipulation of suspended particles in fluidic microsystems is generallyknown and has for example been described by G. Fuhr et al in“Naturwissenschaften”, vol. 81, 1994, p. 528 ff. The microsystems formin particular channel structures through which a suspension fluid flowswith the particles to be manipulated. As a rule the cross-sectional areaof these channel structures is rectangular, with the width of thechannel walls, which in operating position form the top and bottom(bottom/cover surfaces), being greater than the lateral channel walls(lateral surfaces). In the channel structures, microelectrodes areaffixed to the channel walls, with high-frequency electrical fieldsbeing applied to said microelectrodes. Under the influence of thehigh-frequency electrical fields, based on negative or positivedielectrophoresis, polarisation forces are generated in the suspendedparticles, said polarisation forces making possible repulsion from theelectrodes and, acting in combination with flow forces in the suspensionliquid, making possible manipulation of the particles in the channel. Asa rule, the microelectrodes of conventional microsystems are applied asstraight electrode bands to the wider channel walls.

To generate the high-frequency electrical fields effective fordielectrophoresis, in each instance two electrode bands act incombination, said electrode bands being located at opposite channelwalls, both with the same shape and alignment. For example the straightelectrode bands are aligned parallel to the alignment of the channeli.e. the direction of flow of the suspension liquid in the respectivechannel section or at a predetermined angle transversely to thealignment of the channel. For an effective and safe formation of polarisat ion forces at the particles, the length of the electrode bandsexceeds the characteristic dimensions of the particles to be manipulatedmany times over (by a factor of approx. 20 to 50).

Conventional microsystems have disadvantages in relation to theeffectiveness of generating polarisation forces; the stability andlongevity of the microelectrodes; and a limited ability of generatingforce gradients within the channel structure. These disadvantages are inparticular linked to the electrode bands which are formed alongcomparatively long lengths in the channel. The longer an electrode band,the longer a particle flowing past is in the sphere of influence of theelectrode band. Consequently, the effectiveness of the respectivemicroelectrode or the field barrier generated by said microelectrode,increases. However, long electrode bands are also more susceptible tomalfunction. Faults in workmanship or mechanical loads can causeinterruptions which lead to electrode failure. Furthermore, to achieve aforce effect which remains constant along the length of the channel, andthus a reproducible force effect, microelectrodes have so far beenlimited to the above-mentioned straight electrode shape.

Due to the disadvantages mentioned, the application of said fluidicmicrosystems with dielectrophoretic particle manipulation has beenlimited to guiding the particles in the channel structure or todeflecting particles from a given flow.

SUMMARY OF THE INVENTION

It is the object of the invention to provide improved micro systems fordielectrophoretic particle deflection, with said arrangements beingsuitable to overcome the disadvantages of conventional microsystems, andin particular providing enlarged applications and making it possible togenerate field barriers which are also effective in covering shorterchannel sections. Furthermore it is the object of the invention toprovide novel applications for such microsystems.

A microsystem according to the invention is in particular adapted tocreate field barriers in the microsystem along predetermined referencesurfaces, said field barriers extending at least partly across the widthof a channel in the microsystem, and comprising predetermined curvaturesrelative to the longitudinal extension of the channel, to the directionof flow of the suspension liquid in the channel or to the direction ofmovement of the (non-deflected) particles. In this context the term“reference surface” not only describes a two-dimensional formation butalso a spatial region to which the field effect of the respectivemicroelectrodes extends and in which the field barrier for dielectricinfluencing of the microscopic particles in the microsystem is formed.This spatial region essentially corresponds to a region through whichthe field lines of the effective microelectrodes pass; in the case ofmicroelectrode pairs acting in combination, said spatial regionessentially passes as a curved hypersurface between the microelectrodes,while in the case of individually acting microelectrodes it acts as ahypersurface encompassing the field line distribution of theindividually acting microelectrode. Reference surfaces define thelocations where polarisation forces in the microscopic particles caneffectively be generated. The microelectrodes are designed such that thereference surfaces, depending on the desired function of the respectivemicro electrodes, have a predetermined curvature in relation to thedirection of movement of the particles in the microsystem, so that anoptimal combined effect of the polarisation forces and of the mechanicalforces is achieved. Therefore the field barriers are also referred to asfunctional field barriers. The term “curvature” used here does not referto the curvature of field lines on straight microelectrodes as a resultof the field lines exiting into the adjacent space. But rather,“curvature” refers to the shape of the field barriers formed onmicroelectrodes.

Preferably, the field barriers with the reference surfaces curvedaccording to the invention are formed according to one of the followingthree basic forms. According to a first variant, an electrodearrangement according to the invention comprises at least oneband-shaped, curved microelectrode extending on the wider channel wall(bottom surface and/or cover surface), at least partly across thechannel width. In a second variant, at least one microelectrode isprovided which is affixed to the narrower channel wall (lateralsurface). In the third variant, at least one microelectrode is affixedto the bottom surface and/or the cover surface of the channel and atleast one auxiliary electrode is affixed at a distance from the bottomsurface or lateral surface of the channel. The auxiliary electrodesupplies a deformation of the field lines emanating from themicroelectrode or from the microelectrodes at the bottom surfaces or theside surfaces of the channel so that the reference surfaces curvedaccording to the invention, are formed. In all the variants, therespective electrodes (microelectrodes, auxiliary electrodes) per se canbe band-shaped or point-shaped or area-shaped. The electrodearrangements of the second and third variant are also referred to asthree-dimensional electrode arrangements, because microelectrodes areused which protrude from the planes of the bottom surfaces or lateralsurfaces of the channel or which are arranged at a distance from saidsurfaces.

It is thus a subject of the invention, to optimise microelectrodes inrelation to their effect on suspended particles which may comprisenatural or synthetic particles, e.g. for generating maximum forces whileat the same time causing minimum electrical losses.

The invention provides the following advantages. The design of themicroelectrodes can e.g. be adapted to the flow profile in thesuspension liquid. This provides the advantage that the microelectrodescan be shorter and can be designed for generating lesser barriers whilebeing equally as effective as conventional microelectrodes in the shapeof straight bands. This has an advantageous effect on the lifetime andfunction of the microelectrodes and thus of the entire microsystems.Moreover the space available in a microsystem can be used moreeffectively. Furthermore, electrode arrangements are provided with whichgradients can be generated, and thus depending on the respective channelregion, forces of various strength can be generated. It is for exampleprovided for the field barriers of the microelectrodes to be designedsuch that larger polarisation forces act upon the particles in themiddle of the channel when compared to the particles at the edge of thechannel.

The creation of field barriers according to the invention along curvedreference surfaces also makes it possible to create novel applicationsof microsystems, in particular for guiding suspended particles intoparticular channel regions, for sorting suspended particles according totheir passive electrical properties or for collecting or retainingsuspended particles in particular channel sections. For this latterapplication, the microelectrodes are designed so as to comprise ageometric shape for retaining particles in a solution flow or forgenerating a particle formation. All the applications mentioned providecontactless manipulation of the suspended particles vis-a-vis themicrosystem, a feature which is significant in particular formanipulating biological cells or cell components.

Preferred applications are in the field of microsystem technology forseparation, manipulation, loading, fusion, permeation, pair formationand aggregate formation of microscopic small particles.

According to a particular embodiment of the invention, particle movementtakes place in a microsystem with conventional electrode shapes orelectrode shapes according to the invention, under the influence ofcentrifugal forces and/or gravitational forces.

BRIEF DESCRIPTION OF THE DRAWINGS

Further details and advantages of the invention are provided in thedrawings which are described below. The following are shown:

FIGS. 1 a to 1 d illustrates a diagrammatic perspective views of achannel structure with microelectrodes for generating field barriers ina microchannel and examples of reference surfaces curved according tothe invention;

FIG. 2 illustrates a diagrammatic top view of band-shaped curvedmicroelectrodes;

FIG. 3 illustrates a diagrammatic top view of a modified design ofband-shaped curved microelectrodes;

FIGS. 4 a to 4 c illustrates diagrammatic views for illustrating sortingelectrodes for particle sorting;

FIGS. 5 a to 5 b illustrates diagrammatic views of microelectrodes forgenerating field gradients;

FIGS. 6 a to 6 e illustrates diagrammatic views of band-shapedcollecting electrodes according to the invention;

FIGS. 7 a to 7 c illustrates further embodiments of collectingelectrodes according to the invention;

FIG. 8 illustrates a top view of various electrode arrangements forgenerating curved field barriers;

FIG. 9 is corresponding to FIG. 1 d;

FIG. 10 illustrates a diagrammatic top view of a segmented electrodearrangement; and

DETAILED DESCRIPTION OF THE INVENTION

FIG. 1 a diagrammatically shows an example of an embodiment ofmicroelectrodes for generating field barriers in microchannels. Thefluidic microsystem 20 is shown in sections in distorted perspectivelateral view of a channel structure. The channel 21 is formed by twospacers 23 arranged at a distance on a substrate 22, spacers 23supporting a cover part 24. The width and height of the channel isapprox. 200 pm and 40 pm respectively but they can also be less. Suchstructures are for example made using process techniques ofsemiconductor technology which are known per see The substrate 22 formsthe bottom surface 21 a of the channel 21. Accordingly the cover surface(for reasons of clarity not specially emphasised) is formed by the coverpart 24. The electrode arrangement 10 comprises microelectrodes 11, 12attached to the bottom surface 21 a or the cover surface. Each of themicroelectrodes 11, 12 comprises curved electrode bands which aredescribed in more detail below.

In FIG. 1 a the electrode bands form an electrode structure which isexplained in detail below, with reference to FIG. 2. The otherembodiments, described below, of electrode arrangement according to theinvention, can be affixed to the bottom, cover and/or lateral surfacesof the channel 21. A suspension liquid flows through the microchannel 21(from left to right in the illustration), with particles 30 beingsuspended in said suspension liquid. For example, it is the task of theelectrode arrangement 10 shown in FIG. 1 a, to lead the particles 30from various tracks of movement within the channel to a middle track ofmovement according to arrow A. To this effect electrical potential isapplied to the microelectrodes 11, 12 such that electrical fieldbarriers are generated in the channel which force the particles flowingfrom the right, to move to the middle of the channel (direction ofarrows B).

Typical dimensions of the microelectrodes 11, 12 are 0.1 to some tens ofmicrometers (typically 5 . . . 10 pm) in width, 100 nm to a fewmicrometers (typically 200 nm) in thickness, and up to several hundredmicrometers in length. Due to the small thickness of the electrodes, theinterior of the channel 21 is not restricted by the top and bottom ofthe parts 23, 24 of the electrodes processed. Part 23 is a spacer whosestructure forms the lateral channel walls.

The microelectrodes 11, 12 are selected by means of high-frequencyelectrical signals (typically at a frequency in the MHz range and at anamplitude in the volt range). The respective opposite electrodes 11 a,11 b form a control pair although also the electrodes aligned in oneplane can combine the effect of their selection action (phase,frequency, amplitude). The electrical high-frequency field generatedthrough the channel 21, i.e. perpendicular to the direction of flow,acts in a polarising way on the suspended particles 30 (which can alsobe living cells or viruses). At the frequencies mentioned and withsuitable conductivity of the suspension liquid surrounding theparticles, the particles are repulsed by the electrodes. In this way thehydrodynamically open channel 21 can be structured via the electricalfields with a switch-on and switch-off action, or compartmentalised, orthe tracks of movement of the particles in the passive flow field can beinfluenced. Furthermore, it is possible, despite the permanent flow, toslow down the particles or to position them on station without touchinga surface. The type and implementation of the electrode arrangementformed for this purpose also forms part of the invention.

Below, different forms of electrode arrangements according to theinvention are described. For reasons of clarity, FIGS. 2 to 13 may onlyshow a planar electrode arrangement (or parts thereof), e.g. on thebottom surface of the channel.

FIGS. 1 b to 1 c show the basic forms of field barriers orelectromagnetic limitations which are implemented with electrodearrangements according to the invention corresponding to theabove-mentioned variants. The illustrations are schematic diagrams ofthe reference surfaces on which the field barriers are formed withmicroelectrodes according to the invention. For the sake of clarity,only parts of the lateral surface (spacers 23) and of the bottom surface21 a of the channel, the microelectrodes 11, 12 and the shape of thereference surfaces (hatched), are shown.

According to the above-mentioned first variant, the field barrier in thechannel is formed between two curved microelectrodes 11, 12 on thebottom or cover surfaces of the channel (FIG. 1 b). Accordingly, thereference surface of the field barrier (shown hatched) is a curvedsurface positioned vertically against the bottom and cover surfaces. Ifthe microelectrodes 11, 12 are for example curved according to aparticular hyperbolic flow profile (see below), then the referencesurface forms the cutout of the generated surface of a hyperboliccylinder. If the microelectrodes 11, 12 are not arranged exactly on topof each other, then the reference surface is also oblique-angled inrelation to the bottom and cover surfaces of the channel.

According to FIG. 1 c, the reference surface, shown hatched, shows aspatial region impinged on by field lines extending from onemicroelectrode 11 at a lateral surface of the channel to amicroelectrode 12 at the opposite lateral surface. In the example shown,the surface of the first microelectrode 11 is larger than that of thesecond microelectrode 12 so that there is a field line concentration atmicroelectrode 12. Consequently, the polarisation forces acting from thefield barrier on suspended particles are larger near the secondmicroelectrode 12 than near the first microelectrode 11 (see also FIG.9).

FIG. 1 d shows the above-mentioned third variant with athree-dimensional electrode arrangement. The microelectrodes 11, 12 areon the bottom or cover surfaces of the channel while the auxiliaryelectrode 13 with a suitable confinement is arranged in the middle ofthe channel (see also FIG. 10). As a result of the auxiliary electrode13, the field lines between the microelectrodes 11, 12 are distorted,resulting in the curved reference surface, shown hatched (shown in partonly).

The illustrated reference surfaces only represent the positions of thefield barriers without illustrating the forces acting in the respectiveregions, i.e. the height of the field barriers. Essentially the actingforces depend on the density of the field lines and the passiveelectrical characteristics of the particles to be manipulated in therespective channel region. The functional field barriers according tothe invention are thus influenced by the geometric shape of themicroelectrodes which combine their effect, both in relation to theirshape (curvatures etc.) because the dielectrophoretic repulsion forcesare essentially positioned perpendicular to the reference surfaces, andin relation to their areas (field line density).

FIG. 2 shows an electrode arrangement 10 according to the inventionaccording to the above-mentioned first variant. Microelectrodes 11 a, 11b are arranged on the bottom surface 21 a of the channel of amicrosystem, said channel being laterally delimited by the spacers 23.High-frequency electrical potential is applied to the microelectrodes 11a, 11 b via the control lines 14; said microelectrodes 11 a, 11 b act incombination to form a so-called particle funnel as follows.

The electrode arrangement 10 is intended to touchlessly focus, onto themiddle line of the channel, the particles 30 a flowing along the entirewidth of the channel or the entire volume, as is shown by the positionof particle 30 b. This arrangement has the advantage of optimising theelectrode bands in relation to ensuring deflection (focussing) of thesuspended particles, a shortening of the electrode arrangement inlongitudinal direction of the channel and a reduction in electricallosses at the microelectrodes.

In this embodiment of the invention, the basic idea of the design of themicroelectrodes consists of adapting the curvature of the referencesurfaces formed by the field barrier, to the flow forces in the channel.For in microsystems with channel dimensions below 500 pm, due to theReynolds number being low at these dimensions, laminar flows withpredefined flow profiles form. The flow speed near the channel walls islower than that in the middle of the channel (flow speed directly at thechannel wall equals zero). As a result, the flow forces occurring nearthe channel walls are less than those in the middle of the channel. Thismakes it possible to manipulate the particles at the edge of the channelwith lesser polarisation forces or with polarisation forces more steeplydirected against the flow forces than in the middle of the channel. Thecombined effect of the flow forces and polarisation forces is explainedbelow. If essentially the same polarisation forces are formed along theentire length of the microelectrodes, it is sufficient for safedeflection, for the particles to be manipulated, at the edge of thechannel to encounter microelectrodes protruding more steeply into thechannel than they do in the middle of the channel. This makes itpossible to achieve a significant shortening of the microelectrodes (seebelow)

In FIG. 2 the forces acting on the particles are illustrated as anexample in individual sections of the microelectrode 11 a. Therespective total force is composed from the electrically-inducedrepulsion force Fp (polarisation force) and the driving force Fs whichis exerted by the flow of the suspension liquid or from the exterior(e.g. in centrifugal systems as centrifugal force). The resulting totalforce FR results from vector addition of forces Fp and Fs. If the vectorof the total forces FR does not intersect the field barrier of themicroelectrode 11 a, then a particle is safely deflected. The forcediagrams in FIG. 2 illustrate that the driving force Fs increasestowards the middle of the channel. To meet the above-mentioned conditionof safe particle deflection, accordingly the angle between the alignmentof the microelectrode 11 a and the longitudinal direction of the channelchanges from a steeper angle at the edge of the channel to a shallowangle (near-parallelity) in the middle of the channel.

Thus the microelectrodes 11 a, 11 b are curved depending on the flowprofile. In the embodiment shown, each of the band-shapedmicroelectrodes consists of a multitude of straight electrode sections.But in a variation of this embodiment, a steady curvature can beprovided. Corresponding to the parabolic or hyperbolic flow profileoccurring in laminar flows, the curvature is also parabolic orhyperbolic.

According to the invention, the microelectrodes 11 a, 11 b form thefield barriers along a curved reference surface.

The microelectrodes 11 c, 11 d are not provided for in practicalapplication; in the illustration they serve the purpose of comparing anarrangement according to the invention of polygonally curvedmicroelectrodes with straight electrode bands providing the samedeflection performance. It has been found that the microelectrodes 11 a,11 b according to the invention are considerably shorter.

The narrow electrode bands shown in FIG. 2 are very sensitive to faultsin workmanship and local interruptions. A hairline crack at the shoulderof a band-shaped microelectrode leads to failure of the entiremicroelectrode. This can be overcome by an electrode design as showndiagrammatically in FIG. 3. The structuring and covering techniquedescribed in relation to FIG. 3 can also be implemented with otherembodiments of the invention.

FIG. 3 shows a microelectrode 11 with a control line 14. The electrode11 consists of an electrically conductive layer 15 which carries anelectrically non-conductive insulation layer or cover layer 16. Theinsulation layer 16 comprises structured recesses which expose the layer15. In FIG. 3 the insulation layer 16 is shown hatched, while the (e.g.metallic) layer 15 is shown in black. Structuring of the insulationlayer takes place according to the desired form of microelectrodes whichin the example shown are arranged to form a particle funnel as shown inFIG. 2. The electrical field lines emanate from the metallic layer 15into the channel only in the regions of the recesses, so that againfield barriers with reference surfaces that are curved in anapplication-specific way are formed. This arrangement has the advantagethat a small interruption of the exposed sections of the metallic layer15 (i.e. of the microelectrode) does not lead to failure because therespective potential is also applied to the remaining metallic layer 15.For example the thickness of the layer 15 is approx. 50 nm to severalpm, typically approx. 200 nm. The thickness of the insulation layer isaround 100 nm to several pm. Preferably the insulation layer comprisesbiocompatible materials (e.g. oxides, Si02, SiN03 and the like,polymers, tantalum compounds or the like).

Below, a further embodiment of the electrode arrangement 10 according tothe above-mentioned first variant is explained with reference to FIGS. 4a to 4 c. An important application of fluidic microsystems consists ofsorting the suspended particles depending on their passive electricalcharacteristics (hereafter referred to as polarisation characteristicsduring negative dielectrophoresis). Polarisation characteristics dependon the dielectric properties of the particles and their dimensions. Thedielectric characteristics of biological cells are a sensitive indicatorof certain cell characteristics or cell changes which per se could notbe detected for example by monitoring cell size.

Sorting of particles depending on their passive electricalcharacteristics is based on the following principle. Whether or not aparticle can pass the field barrier formed by a sorting electrodedepends on whether or not the resulting force from driving force Fs andpolarising force Fp (see above) intersects the field barrier. If theresulting total force FR points through the field barrier, then theparticle moves in this direction, i.e. the sorting electrode is passed.However if the resulting force FR points to a region located upstream inrelation to the sorting electrode, then the particle will move in thisdirection and will not be able to pass the sorting electrode. Asexplained above, the resulting force FR depends on the flow speed in thechannel and thus the x position of the particles. The flow speedincreases towards the middle of the channel. Thus particles ofrelatively high polarisability which cannot pass the sorting electrodeat the edge of the channel, are subjected to a stronger driving force Fstowards the middle of the channel, so that they may then possibly movepast the sorting electrode. The change in flow speed in x-directionfollows the flow profile; as a rule it is non-linear. Accordingly, theuse of a straight sorting electrode would result in non-linearseparation behaviour. The implementation of curved field barriersaccording to the invention compensates for this. To this effect,microelectrodes 41 a, 41 b with a curvature depending on the flowprofile are used according to the principles explained with reference toFIG. 2.

FIG. 4 a shows two examples of curved microelectrodes 41 a, 41 b on thebottom surface 21 a of a channel between lateral spacers 23. The flow inthe channel is in y direction from left to right, with the arrows vshowing the speed-flow profile in the channel. Upstream, in front of theactual sorting electrode 41 a or 41 b there is a straight microelectrode47 whose task is to focus to a start line s, the particles 30 flowing infrom the left. The microelectrode 47 can also be designated a focussingelectrode. It can be a straight conventional deflector electrode (asshown) or a curved deflector electrode. Downstream of the focussingelectrode 47, one of the sorting electrodes 41 a or 41 b is arrangedwhose task consists of leading the inflowing particles 30 into differenttracks in x-direction in the channel, depending on their polarisationcharacteristics. Particles with a high polarisability should move onwardin different tracks in y-direction than particles with a lowpolarisability 30 b.

The sorting electrode 41 a is set up for linear force effect. For thispurpose the curvature of the microelectrode is shaped according to theflow profile. If the flow speed is low, the setting angle between themicroelectrode and the y-direction is steep; if the flow speed is high,the setting angle is more shallow. Thus the microelectrode 41 a isS-shaped with a turning point in the middle of the channel. After aparticle has passed the sorting electrode 41 a there is a linearrelationship between the x-co-ordinate of the particle and itspolarisability. If a non-linear sorting effect is intended, then themicroelectrode can be curved like the sorting electrode 11 b. Thecurvature is less pronounced than is the case with sorting electrode 11a so that the influence of the driving force Fs is not compensated forby the flow speed. Depending on the relationships set, a non-linearinfluence arises between the x-position of the particles and theirpolarisability after passing the sorting electrode 11 b. Thisconfiguration can in particular be used to separate two particle typesof different polarisability.

Results of experiments show that with a sorting arrangement according toFIG. 4 a, it was possible to neatly separate erythrocytes from so-calledJurkart cells, although both cell types were of the same size.

If the flow profile in the channel is not distinctly parabolic in shapeas shown in FIG. 4 a, but instead is plateau shaped, then sortingelectrodes 41 c, 41 d according to FIG. 4 b are provided. The flow speedfirst increases from the direction of the edge of the channel beforeremaining essentially constant in the middle section of the channel. Soas to achieve a linear sorting effect, the band shape of sortingelectrode 41 a is straight, while at its ends there are curvatures totake into account the changing driving force Fs. For a non-linearsorting effect, the sorting electrode 41 d is curved. From the shoulderof the sorting electrode 41 d at the control connection 14 to its endthere is an increasing effect of the field barrier.

The shape of the sorting electrodes can also be adapted to morecomplicated flow profiles as is shown in FIG. 4 c. In the microsystem 20a first channel 211 with a high flow speed and a second channel 212 witha lesser flow speed join to form a joint channel 21. Due to thelaminarity of the flow, the flow profile at first remains intact also inthe common flow path. Accordingly, the sorting electrodes 41 e or 41 fare curved so as to achieve a particular linear or non-linear sortingeffect. The lower the flow speed, the higher the setting angle betweenthe direction of the microelectrode (alignment of the reference surface)and the longitudinal direction of the channel (y-direction).

For reasons of clarity, the focussing electrode 47 according to FIG. 4a, is not shown in FIGS. 4 b and 4 c.

The above-mentioned sorting takes place with the assumption that thepotential is constant along the entire length of the microelectrode. Inreality however, small electrical losses occur along the microelectrodeso that the field barrier becomes progressively smaller from theshoulder of the microelectrode (at the control line) towards its end.This phenomenon can be taken into account in the curvature of thesorting electrodes in that at the side of the control line of thechannel, a larger electrode curvature is provided than at the end of thesorting electrodes. But the phenomenon mentioned can also bepurposefully used for additional non-linear separation effects. By usingmodified embodiments, the loss of potential towards the end of themicroelectrodes can be amplified especially as a result of measures forforming field gradients. This means that the height of the field barrierformed by the microelectrode increases or decreases along the curvedelectrode band. Such gradient electrodes can be designed in a shapeaccording to FIG. 5.

For particle sorting in relation to various groups of characteristics,several sorting electrodes according to FIG. 4 can be arranged insequence in the direction of the channel. A characteristic potential orpotential gradient at a predetermined frequency is applied to eachsorting electrode. For example relatively low frequencies (in the regionof approx. 10 kHz) can be used for sorting in relation to variousdielectric membrane characteristics, and high frequencies (above 100kHz) can be used for sorting depending on the cytoplasmatic conductivityof biological cells.

FIG. 5 shows gradient electrodes 51 a, 51 b; for the sake of clarity,the electrode bands are straight. To set field barriers according to theinvention, with curved surfaces, the gradient electrodes shownadditionally comprise a characteristic application-specific curvatureaccording to the principles explained above.

The gradient electrode 51 a is formed by a closed electrode band arounda triangular surface. As the distance from the control line 14increases, the field line density is reduced in line with the fanningout of the triangle. The same applies to the gradient electrode 51 bwith two diverging partial bands 511 b and 512 b.

The collection and at least temporary arrangement of particles orparticle groups in the channel through which suspension liquid flows, isa further important application of fluidic microsystems. To this effect,electrode arrangements according to the invention are shaped ascollecting electrodes as is explained below with reference to FIGS. 6 to8.

FIG. 6 a shows the basic shape of a collecting electrode. Again, onlyone microelectrode on the bottom or cover surface of a channel is shown,which acts in combination with that of a second microelectrode locatedat the opposite side of the channel. A collecting electrode 61 acomprises an electrode band with an angle section 611 a and a supplysection 612 a. The angle section 611 a forms an angle pointing in thedirection of flow (x-direction). The opening angle of the angle section611 a is selected depending on the shape of the particles to becollected; preferably the opening angle is less than 90°, e.g. rangingfrom 20 to 60°. The opposing angle sections of electrodes whose effectis combined, form a barrier which the particles 30 to be collectedcannot pass despite the driving force of the flow. This barrier ismaintained as long as the collecting electrodes remain selected. Thesupply section 612 a is electrically ineffective as a result of aninsulation layer 16. FIG. 6 b shows a modified embodiment of acollecting electrode 61 b which is made with reference to the covertechnique explained above with reference to FIG. 3. The electricallyeffective angle section 611 b is formed by a recess in the insulationlayer 16 as a result of which a deeper metallic layer 15 is exposedtowards the suspension liquid containing the particles.

FIGS. 6 c and 6 d show corresponding collecting electrodes 61 c and 61d, each comprising a multitude of angle sections 611 c or 611 d. Theseangle sections are again set to collect inflowing particles 30. Byadjoining the angle sections 611 c or 611 d across the longitudinaldirection (x-direction) of the channel, the particles flowing in thevarious channel sections can be collected selectively. A collectingelectrode 61 c or 61 d is advantageously combined with one of thesorting electrodes according to FIGS. 4 a to 4 c. The sorted particlesare separately collected in the individual collection regions of thecollection electrodes. The collection electrode 61 d essentiallycorresponds to the collection electrode 61 c. This completesimplementation of the entire cover technique.

The collection electrodes 61 c or 61 d are particularly well suited toline up particles in the suspension flow in the manner of a start linefrom which the particles flow onward simultaneously when the controlpotential of the collection electrodes is switched off.

FIG. 6 e shows a further embodiment of a collection electrode 61 e wherea multitude of angle sections 611 e is also provided, but with the anglesections being designed for collecting or gathering particles of varioussizes or various sized accumulations of such particles.

FIG. 7 a shows the accumulation of a particle group 300 using acollecting electrode 71 a. This embodiment of a collecting electrodediffers from the collecting electrode according to FIG. 6 a only in itsdimensions. This design is particularly well suited to the formation ofparticle aggregates. Again, preferably a combination with a sortingarrangement according to FIGS. 4 a to 4 c is implemented.

The electrode arrangement according to FIG. 7 b is configured forseparate collection of particles or particle groups from the suspensionflow in the channel which differ in relation to their flow track inx-direction. The microelectrode arrangement 71 b comprises severalpartial collection electrodes each with an angle section 711 b, wherebyeach partial collection electrode can be selected separately. Whencombining such a collecting electrode arrangement with a sortingarrangement according to FIGS. 4 a to 4 c, the following method can beimplemented with particular advantage.

First a particle mixture which flows through the channel in themicrosystem is sorted depending on the passive electricalcharacteristics of the particles, and in this way is guided to varioustracks mutually spaced apart in x-direction. Then in a collectionelectrode according to FIG. 7 b, particle-type specific collection ofthe particles flowing in the individual tracks takes place. By releasingthe partial collection electrodes according to a time sequence (in eachinstance by switching off the control potential), the previously sortedparticles can flow on in groups in the microsystem. Downstream, thechannel can for example be split into several sub-channels, with thegroups of particle types specifically being directed to saidsub-channels.

FIG. 7 c shows a further collecting electrode 71 c for generating apredetermined particle formation.

Depending on the application, the angle sections of the collectingelectrodes shown in FIGS. 6 and 7 can extend across the entire width ofthe channel or only across parts of the channel. Within an electrodearrangement, collecting electrodes can be provided for individualparticles and/or for particle groups.

Further embodiments of combined sorting electrodes and collectionelectrodes are shown in FIG. 8 in top view of the bottom surface 21 a ofa channel delimited by spacers 23. The suspension liquid with suspendedparticles flows through the channel in y-direction. According to FIG. 8a an area-shaped microelectrode 81 a on the bottom surface 21 a and astraight band-shaped microelectrode 82 a (shown by a dashed line) on theopposite surface of the channel, act in combination. The planar-shapedmicroelectrode 81 a has been produced using the cover techniqueexplained above. A metallic layer supports an insulation layer with arecess according to the shape of the microelectrode 81 a (drawn inblack). The field lines between the microelectrodes 81 a and 82 a areinhomogenously aligned across the direction of flow, resulting in anasymmetric field barrier or again a reference surface which is curvedaccording to the invention. In the middle of the channel the field linedensity is largest so that the electrically generated forces are locatedin the region of the highest flow speed. In this way an essentiallyconstant balance between the driving force resulting from the flow andthe electrical polarisation force is formed in x-direction across thewidth of the channel. According to FIG. 8 b, again a field barrier witha curved reference surface is formed. The microelectrodes 81 b, 82 b areboth designed so as to be linear or band-shaped, they are not arrangedin opposite positions but instead offset in relation to each other.

FIG. 8 c shows an electrode arrangement for forming particle aggregates.The microelectrodes 81 c, 82 c form a number of funnel-shaped particlecollectors arranged side by side. Each particle collector 81 is formedby a field barrier which in the direction of the flow of liquid firstnarrows in the form of a funnel before discharging into a straightchannel section 812. The channel section is dimensioned such that twoparticles can be arranged one behind the other in the direction of flow.Due to the formation of adhesion forces, the particles form an aggregate(so called pair-loading in the direction of flow). The embodimentaccording to FIG. 8 d is a modified version in that pair-loading takesplace across the direction of flow. The individual collector elements811 d comprise electrode tips 813 d on the inlet side. With theseelectrode tips 813 d an additional barrier effect or filter effect canbe achieved, and already existing aggregates or larger particles 30 dcan be precluded from assembling in the collecting electrode 81 d.

FIG. 9 (corresponding to FIG. 1 d) shows an embodiment of an electrodearrangement according to the above-mentioned third variant. In amicrosystem, again two sub-channels 211, 212 extend parallel to eachother and separated from each other by a separation wall 231 comprisingan aperture 232. The electrode arrangement according to the inventioncomprises microelectrodes on the bottom surfaces and cover surfaces inthe form of focussing electrodes 101, 102 and the auxiliary electrode103. The auxiliary electrode is arranged at the separation wall 231,adjacent to the aperture 232, on the downstream side of the aperture232. The auxiliary electrode 103 does not comprise a control line. Itmerely serves to form the reference surface of the field barriers formedby the electrode arrangement. The effect of the microelectrodes iscombined as follows.

Focussing electrodes 101 and 102 are used to focus the particles 30 a,30 b, flowing in the sub-channels 211 or 212, to a middle line accordingto the position of the aperture 232 in the separation wall 231. Theparticles are deflected by the field barrier between he focussingelectrode 101 and the auxiliary electrode 103 or between the focussingelectrode 102 and the auxiliary electrode 103, through the aperture 232,into the adjacent sub-channel or they are left in the respectivesub-channel. According to a preferred method, the focussing electrodes101, 102 are operated at various frequencies so as to act in aparticle-selective manner. Accordingly, again selective particle sortinginto the sub-channels, or deflection of predetermined particles to anadjacent sub-channel can be achieved to carry out a particular treatmentof the active ingredient with the respective suspension liquid providedin said sub-channel.

According to a particular aspect of the invention, the microelectrodesin the individual embodiments can be segmented per see However, in thiscase each microelectrode comprises a number of electrode segments whichare arranged according to the desired electrode function. FIG. 10 showsa particularly versatile microelectrode 131 as an array of a multitudeof pixel-shaped electrode segments arranged matrix-like. The electrodesegments are arranged across the entire width of the bottom surface 21 abetween the spacers 23 and can be selected individually. This makes itpossible to form the desired curved field barriers in particularaccording to the above-mentioned first variant, depending on theconcrete application, in particular depending on the particles to bemanipulated, the flow conditions and the task of the microsystem. InFIG. 13 the presently selected pixels are shown in black while thepixels which are not selected are shown in white. In this case thesegmented microelectrode 131 assumes the function of a particle funnelaccording to FIG. 2 by means of which the particles 30 are focussed tothe middle of the channel.

The pixel-shaped electrode segments make possible loss-minimisingfocussing, sorting or collecting of particles. Each electrode segmentcan be selected with its own potential value (voltage) or its ownfrequency. In this way any specified dielectric force field can begenerated along the channel. For example the influence of the flowprofile can be compensated for in that the pixels arranged across thelongitudinal direction of the channel are selected with a voltage whichcorresponds to the square root of the profile of the flow speed.

The size of the electrode segments and spacings between the electrodesegments are preferably smaller than the characteristic dimension of theparticles to be manipulated, but they can also be larger.

All particle manipulation takes place contact-free, so that themicrosystems according to the invention are particularly suitable formanipulating biological cells or cell components.

The microsystems are further characterised in that they may compriseapertures (inflows, through-flows, outflows) which can be closed off sothat after or before centrifugation, the particles can easily be removedor inserted. Furthermore, all the microelectrode elements (holdingelectrodes for particles, microfield cages etc.) can be installed whichare known per se for dielectrophoretic influencing of particles, andwhich are used in conventional microsystems which operate with flowingliquids. Based on the combined action of gravitational or centrifugalforces with dielectrophoretic forces, the method according to theinvention is an electrically controlled or active centrifugation.Furthermore, combinations can be provided with the effect of opticalforces (laser tweezers), magnetic forces (influence on magneticparticles), or mechanical forces in the form of ultrasonic forces.

Areas of application of the invention include in particular: cellseparation/cell fractionation, cell sorting, cell loading (molecular,nano-particles, beads), cell discharge (molecular), cell permeation(so-called electroporation), cell fusion (so-called electrofusion), cellpair formation, and cell aggregate formation.

The invention is not limited to particular solution liquids orsuspension liquids. It is advantageous if the viscosity of the liquidcontained in the microsystem is known. If the viscosity is known, therotational speed for setting a particular particle speed can bedetermined on the basis of tabular values or by means of a programalgorithm. Alternatively, it is however also possible to acquire theactual speed of the particles in the microsystem during centrifugation(e.g. by using an optical sensor) and to regulate the rotational speedfor setting a particular particle speed. It can be provided that invarious sub-sections of the channel structures, e.g. in parallelchannels which are interconnected only via an aperture, liquids ofvarious viscosity are contained. In this case however, viscosities arepreferred which ensure that diffusion of the liquids through theaperture is relatively low or negligible over the entire period ofcentrifugation.

If the mass density of the particles is less than that of the liquid inthe microsystem, the invention can be implemented with correspondingmodifications in that particles are introduced on the side of themicrosystem away from the axis of rotation. They then move to the otherend of the microsystem under the influence of buoyancy or by thecombined effect of buoyancy and centrifugal forces.

The microsystem is designed corresponding to the channel structure andalignment of the electrodes in dependence on the particular application.As a rule, the cross-sectional dimensions of channels are significantlylarger than the diameter of individual particles. Advantageously, thisprevents blocking of the channels. If only particles with particularlysmall dimensions have to be manipulated (e.g. bacteria or viruses orcell organelles), then the channel dimensions can be reducedaccordingly, e.g. to dimensions below 10 pm.

The invention is implemented with a microsystem which is closed off atleast on one side. The closed end can be a closed-off end of a channel,a closed-off collection zone or a closed-off hollow space in themicrosystem. With particle manipulation according to the invention,there is essentially no movement of liquid towards the closed end. Inparticular with implementation of collection zones or hollow spaces atthe closed-off end, this means that these, like the entire microsystem,are filled with the solution or suspension for the particles at thebeginning of particle manipulation.

If during manipulation of the particles, agglomerations or temporaryblockages of the channel structures occur, according to the invention itis provided to temporarily increase the rotational speed of thecentrifuge so as to detach the adhering particles and move them on.

1. A microsystem adapted for dielectrophoretic manipulation of particlesin a suspension liquid, said microsystem comprising: a channel withwalls, said channel having a longitudinal extension in an x-directionand said channel walls comprising bottom and cover surface wallsextending in the x-direction and an y-direction, and an electrodearrangement on at least one of said bottom or cover surface walls forgenerating a field barrier which crosses the channel at least partly,wherein said electrode arrangement comprises at least one microelectrodehaving a band-shape, and in relation to the longitudinal extension ofsaid channel, said band-shape has a predetermined parabolic orhyperbolic curvature along its length so that the field barrier has acorresponding parabolic or hyperbolic curvature.
 2. The microsystemaccording to claim 1, in which the electrode arrangement comprises atleast two microelectrodes of the same shape and alignment affixed onopposite channel walls, each of said at least two microelectrodes beingin the shape of a curved band.
 3. The microsystem according to claim 2,in which the at least two microelectrodes depending on a flow profile ofsaid suspension liquid flowing though said channel are curved such thatin every section of a field barrier of the microelectrodes a resultingforce acting on a particle in said suspension liquid points to a regionwhich is situated upstream in relation to the microelectrode.
 4. Themicrosystem according to claim 3, in which the at least twomicroelectrodes comprise four microelectrodes being arranged as focusingelectrodes to form a particle funnel.
 5. The microsystem according toclaim 2, in which the at least two microelectrodes adapted to a flowprofile of said suspension liquid flowing through said channel arecurved such that a resulting force acting on a particle from one end ofeach of the microelectrodes towards the other end describes a change indirection which leads from a region situated upstream to a regionsituated downstream respectively in relation to the at least twomicroelectrodes.
 6. The microsystem according to claim 5, in which theat least two microelectrodes are sorting electrodes providing a fieldbarrier in combination with the flow profile of the suspension liquid inthe channel such that each of the suspended particles with differentpassive electrical characteristics can pass the sorting electrodes on aseparate track depending on the characteristics of said suspendedparticles.
 7. A microsystem adapted for dielectrophoretic manipulationof particles in a suspension liquid, said microsystem comprising: achannel with channel walls, said channel having a longitudinal extensionin an x-direction and said channel walls comprising bottom and coversurfaces walls extending in the x-direction and an y-direction, and anelectrode arrangement with at least one microelectrode on at least oneof said bottom or cover surfaces walls for generating a field barrierwhich crosses the channel at least partly, wherein the at least onemicroelectrode comprises a multitude of straight electrode sectionsconnected with each other, and—in relation to the longitudinal extensionof said channel, said straight electrode sections are arranged withpredetermined, different angles and the field barrier has a parabolic orhyperbolic curvature relative to the x- and y-directions correspondingto the arrangement of said straight electrode sections.
 8. Themicrosystem according to claim 1 or 7, wherein at least twomicroelectrodes of the electrode arrangement are arranged in pairs onthe bottom and cover walls of the channel.
 9. The microsystem accordingto claim 1 or 7, in which at least one microelectrode of the electrodearrangement comprises two microelectrodes being provided on two oppositechannel walls, comprising different geometric shapes.
 10. Method ofdielectrophoretic manipulation of particles in a suspension liquid,using a microsystem according to claim 1 or 7, said method comprisingthe steps of: flowing said suspension liquid through the channel of saidmicrosystem, forming a field barrier with a predetermined curvaturerelative to the direction of flow of said suspension liquid, anddeflecting, sorting, collecting and/or forming microscopic particlesunder the influence of said field barrier.